Biology subjects

Johnston, M. G.

Publications and source records attributed to Johnston, M. G..

6 recordsLinked to original sources

bioRxiv2022

A PDLP-NHL3 complex integrates plasmodesmal immune signaling cascades

The plant immune system relies on the perception of molecules that signal the presence of a microbe threat. This triggers signal transduction that mediates a range of cellular responses via a collection of molecular machinery including receptors, small molecules, and enzymes. One response to pathogen perception is the restriction of cell-to-cell communication by plasmodesmal closure. We previously found that while chitin and flg22 trigger specialized immune signaling cascades in the plasmodesmal plasma membrane, both execute plasmodesmal closure via callose synthesis at the plasmodesmata. Therefore, the signaling pathways ultimately converge at or upstream of callose synthesis. To establish the hierarchy of signaling at plasmodesmata and characterize points of convergence in microbe elicitor-triggered signaling, we profiled the dependence of plasmodesmal responses triggered by different elicitors on a range of plasmodesmal signaling machinery. We identified that, like chitin, flg22 signals via RBOHD to induce plasmodesmal closure. Further, we found that PDLP1, PDLP5 and CALS1 are common to microbe- and SA-triggered responses, identifying PDLPs as a candidate signaling nexus. To understand how PDLPs relay a signal to CALS1, we screened for PDLP5 interactors and found NHL3, which is also required for chitin-, flg22- and SA-triggered plasmodesmal responses and PDLP-mediated activation of callose synthesis. We conclude that a PDLP-NHL3 complex acts as an integrating node of plasmodesmal signaling cascades, transmitting multiple immune signals to activate CALS1 and plasmodesmata closure. Significance StatementPlants close plasmodesmata to restrict cell-to-cell communication after pathogen perception and in response to a range of stresses. All these stimuli trigger callose deposition at the plasmodesmal neck, suggesting a convergence of signaling. We have defined the hierarchy of molecular components and signals required to mediate plasmodesmal closure in immune responses, identifying a PDLP-NHL3 complex as a critical node that integrates multiple signaling cascades to regulate plasmodesmata.

plant biology↗

bioRxiv2022

Comparative phyloproteomics identifies conserved plasmodesmal proteins

Plasmodesmata connect neighbouring plant cells across the cell wall. They are cytosolic bridges, lined by the plasma membrane and traversed by endoplasmic reticulum to connect these cell components between cells and tissues. While plasmodesmata are notoriously difficult to extract, tissue fractionation and proteomic analyses have yielded valuable knowledge of their composition. Most proteomic profiles originate from cell suspension cultures in which simple plasmodesmata dominate and have been exclusively generated from dicotyledonous plant species. Here we have generated two novel proteomes to expand tissue and taxonomic representation of plasmodesmata: one from mature Arabidopsis leaves and one from the moss Physcomitrium patens. We have leveraged these and existing data to perform a comparative analysis that, owing to comparing proteomes from an expanded taxonomic tree, allowed us to identify conserved protein families that are associated with plasmodesmata that likely serve as core structural or functional components. Thus, we identified {beta}-1,3-glucanases, C2 lipid-binding proteins and tetraspanins as core plasmodesmal components, with proteins from P. patens and Arabidopsis maintaining plasmodesmal association across diverse species. Our approach has not only identified elements of a conserved, core plasmodesmal proteome, but also demonstrated the added power offered by comparative analysis. Conserved plasmodesmal proteins establish a basis upon which ancient plasmodesmal function can be further investigated to determine the essential roles these structures play in multicellular organism physiology in the green lineages.

plant biology↗

bioRxiv2022

The Hyaloperanospora arabidopsidis effector HaRxL77 is hypermobile between cells and manipulates host defence

Multicellular organisms require dynamic communication between cells, tissues and organs to integrate responses to external and internal signals. In plants, cell-to-cell communication relies in part on plasmodesmata, which connect adjacent cells and allow the exchange of signals and resources. Upon infection by pathogens, plants act to isolate infected cells from non-infected cells by closing plasmodesmata but pathogens can suppress this defence and maintain plasmodesmata in an open state. To address the question of what a pathogen might gain from keeping plasmodesmata open, we screened effectors from the biotrophic Arabidopsis pathogen Hyaloperonospora arabidopsidis (Hpa) for the ability to move cell-to-cell via plasmodesmata in plant tissues. We quantified the mobility of cytoplasmic effectors and identified six that were hypermobile, i.e., can move further than expected for a protein of that size. Of these, HaRxL77 indirectly modifies plasmodesmatal permeability to facilitate hypermobility and suppresses the flg22-induced ROS burst, suggesting that cell-to-cell mobility of effectors allows defence manipulation ahead of the infection front. Thus, this study provides novel insights into how Hpa exploits plasmodesmata-mediated intercellular connectivity to promote infection, characterising a poorly explored element of plant-pathogen interaction. Author SummaryDuring infection, pathogens secrete effectors into host cells to manipulate them for their benefit. Plant cells are connected via plasmodesmata, and pathogen effectors can use these connections to reach uninfected cells. We asked what a pathogen might gain by having effectors that can travel between cells and thus screened Arabidopsis downy mildew effectors for this capability. We found downy mildew produces many cell-to-cell mobile effectors, and that some of these can move further than we might expect. One hypermobile effector, HaRxL77, can open plasmodesmata to allow this increased mobility, although how it does this is not clear. HaRxL77 interferes with several host defence mechanisms, suggesting that cell- to-cell mobility allows effectors to perturb host defence ahead of the infection front.

plant biology↗

bioRxiv2021

Colletotrichum higginsianum effectors exhibit cell to cell hypermobility in plant tissues and modulate intercellular connectivity amongst a variety of cellular processes

Multicellular organisms exchange information and resources between cells to co-ordinate growth and responses. In plants, plasmodesmata establish cytoplasmic continuity between cells to allow for communication and resource exchange across the cell wall. Some plant pathogens use plasmodesmata as a pathway for both molecular and physical invasion. However, the benefits of molecular invasion (cell-to-cell movement of pathogen effectors) are poorly understood. To begin to investigate this and identify which effectors are cell-to-cell mobile, we performed a live imaging-based screen and identified 15 cell-to-cell mobile effectors of the fungal pathogen Colletotrichum higginsianum. Of these, 6 are "hypermobile", showing cell-to-cell mobility greater than expected for a protein of its size. We further identified 3 effectors that can indirectly modify plasmodesmal aperture. Transcriptional profiling of plants expressing hypermobile effectors implicate them in a variety of processes including senescence, glucosinolate production, cell wall integrity, growth and iron metabolism. However, not all effectors had an independent effect on virulence. This suggests a wide range of benefits to infection gained by the mobility of C. higginsianum effectors that likely interact in a complex way during infection.

plant biology↗

bioRxiv2020

A bootstrap approach is a superior statistical method for the comparison of cell-to-cell movement data

Plasmodesmata are an increasing focus of plant research, and plant physiologists frequently aim to understand the dynamics of intercellular movement and plasmodesmal function. For this, experiments that measure the spread of GFP between cells are commonly performed to indicate whether plasmodesmata are more open or closed in different conditions or in different genotypes. We propose cell-to-cell movement data sets are better analysed by a bootstrap method that tests the null hypothesis that means (or medians) are the same between two conditions, instead of the commonly used Mann-Whitney-Wilcoxon test. We found that that with hypothetical distributions similar to cell-to-cell movement data, the Mann-Whitney-Wilcoxon produces a false positive rate of 17% while the bootstrap method maintains a false positive at the set rate of 5% under the same circumstances. Here we present this finding, as well as our rationale, an explanation of the bootstrap method and an R script for easy use. We have further demonstrated its use on published datasets from independent laboratories.

plant biology↗

bioRxiv2017

Evolutionarily younger and faster diversifying plants are more threatened by extinction

Extinction threatens many species, yet few factors predict this risk across the plant Tree of Life (ToL). Taxon age is one factor that may associate with extinction if occupancy of geographic and adaptive zones varies with time, but evidence for such an association has been equivocal. Age-dependent occupancy can also influence diversification rates and thus extinction risk where new taxa have small range and population sizes. Here we analysed 509 well-sampled genera from across the plant ToL. We found that a greater proportion of species were threatened by extinction in younger and faster-diversifying genera. Repeating our analyses in two large, well-sampled groups, we found that extinction risk increased with evolutionary age in conifer species but not palms. Potential range size decreased in older, non-threatened conifers more strongly than in threatened taxa, suggesting that range size dynamics may explain differing patterns of extinction risk across the ToL with consequences for biodiversity conservation.

evolutionary biology↗